Much is known about the schemes and organization of assembly pathways by which biological structures are formed. Previous research on virus assembly has provided much of this knowledge. Relatively little however is known about the molecular bonds which hold these structures together. The research proposed in this grant application is directed at understanding the molecular mechanism of structure formation in biological systems. The approach is to study the assembly of a single protein species into a complex bacterial virus. Two basic techniques will be used: Genetics and protein chemistry. We propose to isolate many different mutants of Salmonella phage P22 which carry defects in the gene coding for the last protein which assembles into the virus structure. We have designed and tested a clever screening and selection technique to obtain these mutants in large numbers. The protein of interest made by each of these mutants will be unable to assemble onto the virus particle. We propose to genetically map these mutants into the structural gene for the protein using our established set of deletion lysogens and note any fortuitous genetic clustering of this particular phenotype. Each of these assembly defective proteins will be purified and characterized with respect to the amino acid change compared to the wild-type protein. Thus a tabulation of amino acid changes which cause assembly defects will be generated as well as a list of the original wild-type amino acids used in assembly bond formation. Revertants of these mutants will be selected in order to attempt identification of the gene products with which the protein interacts during assembly. Other mutations in the gene for this multifunctional protein will be uncovered by this selection technique. The missing functions of these proteins will be identified, the mutations mapped and the amino acid changes catalogued. The change in viral infectivity will be correlated with the loss of protein functions. Therefore we propose to identify those types of amino acids which are used in the bonds formed in assembly reactions through genetic mutational studies. In addition we will identify those regions of the assembly protein involved in assembly and other functions.